Implementation of a Short-term Treatment Protocol in Anemic Patients before Cardiac Surgery.

BACKGROUND: We assessed whether implementation of an immediate preoperative treatment in anemic patients could result in fewer perioperative packed red blood cell (PRBC) transfusions and improved outcomes in a real-world setting. METHODS: From January 1, 2020, to November 31, 2022, we implemented a perioperative protocol for anemic patients (hemoglobin (Hb) level in women <11.5 g/dL, men <12.5 g/dL), which included subcutaneous erythropoietin α, intravenous Iron, and intramuscular vitamin B12 (all given preoperatively) and per os iron and folic acid given once a day postoperatively. We retrospectively compared all patients receiving the protocol to all eligible patients who were operated upon in the 4 years prior to implementation of the protocol. Primary outcome was amount of PRBC transfusions during surgery and index admission. RESULTS: In the months after protocol implementation, 114 patients who received the treatment protocol were compared with 236 anemic patients in the 4 years prior to who did not receive the protocol. The treatment reduced total PRBC use (control group median 4 [2-7] units vs. treatment 2 [1-3] units, p < 0.0001) and the incidence of postoperative blood products transfusions (treatment group 58 patients, 50.88% vs. control group 177 patients, 75%, p < 0.0001). Hb prior to discharge was higher among the protocol group (treatment median 9 g/dL [8.3-9.5 g/dL] vs. control 8.6 g/dL [8.1-9.1 g/dL], p = 0.0081). CONCLUSION: Despite some differences compared with previously described protocols, the implementation of a perioperative treatment protocol for anemic patients was associated with a reduction in PRBC transfusion in a real-world setting.

Prevalence and Genotype Distribution of HPV Types in Women at Risk for Cervical Neoplasia in Israel.

BACKGROUND: Invasive cervical cancer is caused by human papillomavirus (HPV). OBJECTIVES: To describe the prevalence and genotype distribution of HPV types in women at risk for cervical neoplasia. METHODS: Our study summarized HPV types detected in 6654 samples that were sent to the serology laboratory from cervical clinics in northern Israel between 2006-2014. The HPV test was performed during investigation of atypical squamous cells of undetermined significance (ASCUS) results on Pap tests or due to complaints suggestive of cervical neoplasia. HPV types were classified as high risk (HPV-HR) and low risk (HPV-LR). RESULTS: Of the samples, 46.4% (3085/6654) were HPV-HR positive. Of women with cervical intraepithelial neoplasia 2-3 (CIN 2-3) or cancer, 292/318 (91.8%) and 137/145 (94.5%), respectively, were HPV-HR positive. HPV 16 and HPV 18 were detected in 11.8% of the total samples and in 48.2% and 64.9% of the women with CIN 2-3 and with cancer, respectively. HPV was negative in 8/145 (5.5%) and 26/318 (8.2%) of women with cervical cancer and CIN 2-3, respectively. CONCLUSIONS: This study shows the prevalence of HPV types in women at risk for cervical neoplasia. The sensitivity of all HPV types for CIN 2-3 and cervical cancer was 91.8% and 94.5%, respectively; and of HPV-HR types, 89% and 92.4%, respectively. Triage of HPV-HR types should be considered in women with ASCUS because HPV-HR types were discovered in only 36.7%. The distribution of HPV types in our population is similar to that reported for other developed countries.

Asymptomatic carriage of Pneumocystis jirovecii and cytomegalovirus in lungs of immunocompetent patients.

PURPOSE: Pneumocystis jirovecii (PCP) and cytomegalovirus (CMV) are opportunistic pathogens which cause lung infection in immunocompromised individuals. However, scarce data are available regarding the carriage of CMV or PCP in immunocompetent, non critically ill patients. The purpose of this study was to evaluate the prevalence of PCP and CMV in broncholaveolar lavage of adult immunocompetent, non critically ill patients. METHODS: BAL fluids from immunocompetent patients who underwent bronchoscopy were analyzed by polymerase chain reaction (PCR) for CMV and PCP DNA. We tested CMV antibodies in serum. In patients with positive CMV DNA in lavage fluid, we further analyzed peripheral blood for the presence of CMV DNA. RESULTS: Ninety three patients were included. We did not detect PCP DNA in BAL in any patient. CMV DNA was found in BAL of 5 of 86 CMV IgG positive patients (5.8 %). Patients who were positive for CMV did not differ from patients with negative PCR for CMV regarding demographic and clinical features. CONCLUSION: We did not find PCP colonization in our cohort of patients. However, we found significant prevalence of CMV DNA in BAL from immunocompetent patients, with no evidence of acute CMV infection. This finding may represent colonization by CMV in immunocompetent, non-critically ill individuals.

Human parechovirus type 3 central nervous system infections in Israeli infants.

INTRODUCTION: Human parechoviruses (HPeV) have been recognized as the causative agents of central nervous system (CNS) infection of infants and young children in different parts of the world. The role of HPeV in CNS infection of Israeli infants and children is unknown. OBJECTIVES: To assess the detection rate of HPeV in enterovirus RT-PCR-negative cerebrospinal fluid (CSF) samples obtained during the years 2007-2009 from children 0-5 years old with suspected CNS infection or from very young infants with unexplained fever in four medical centers in Israel. STUDY DESIGN: A total of 367 CSF samples were retrospectively tested for the presence of HPeV RNA using nested RT-PCR assay. Positive samples were further typed on the basis of molecular sequencing. Retrospective analysis of the medical charts was performed. RESULTS: HPeV3 RNA was detected in CSF obtained between May and September 2008 in 13 patients, all of whom were <3 months old (3.5% of all CSFs; 11.3% of all infants<3 months in 2008). The HPeV-positive CSF samples were without pleocytosis. All HPeV3-positive patients recovered without obvious short term sequelae. CONCLUSION: HPeV infection could play an important role in summertime febrile/CNS illness in young infants during specific years with high HPeV activity. PCR detection of parechoviral RNA in CSF should be included in the diagnostic evaluation of fever or CNS infection of neonates and very young infants. The rapid identification of HPeV in CSF could curtail unnecessary empirical antibiotic treatment and shorten hospital stay in selected patients.

[Human papillomavirus types in women with cervical cancer in Haifa District].

BACKGROUND: Human papillomavirus (HPV) is recognized as the absolute cause of cervical cancer and is found in 99% of the Lesions. HPV 16 and 18 are detected in 70% of the cases. Two vaccines against HPV 16 and 18 were approved for use in Israel in recent years. PURPOSE: To determine the prevalence of human papillomavirus (HPV) in women with invasive cervical cancer in the Haifa district, and to see if the vaccine is suitable for our population. METHODS: The study population included 65 women from the Haifa District who were diagnosed with invasive cervical cancer in the Cervix Clinic of Carmel Medical Center. Samples for HPV typing were obtained during the evaluation of those patients. DNA was extracted from brush samples and HPV genotype was determined by nested-PCR followed by sequencing. RESULTS: Out of 65 patients with cervical carcinoma, 64 were found positive for HPV. The main HPV types in our patients were HPV 16 in 53.8% of the patients, HPV 18 in 12.3% of the patients and HPV type 45 in 13.8% of the patients. HPV type 33 was found in 4.6% of the patients, and HPV types 31 and 66 in 3.1% of the patients. Each of HPV types 54, 56, 58 and 59 were found in one patient. The main complaint was postmenopausal bleeding or menometrorrhagia in 24 patients (36.9%), post coital bleeding in 18 patients (27.7%) and 14 patients (21.5%) were evaluated due to an abnormal cytological smear Squamous cell carcinoma was diagnosed in 83% of the patients and cervical adenocarcinoma in 15.1% of them. CONCLUSIONS: The prevalence of HPV types in Haifa district is similar to world prevalence of HPV's, where HPV 16 and 18 cause 66.1% of cervical cancer, while in our study HPV45 and HPV 66 were found in higher proportions of cases than reported worldwide. In our population the vaccine against HPV 16/18 can prevent almost 70% of cases of cervical cancer, but a multicenter study should be performed in order to obtain larger numbers.

Paraoxonase 1 deficiency in mice is associated with reduced steroid biosynthesis: effects on HDL binding, cholesteryl ester accumulation and scavenger receptor type BI expression.

OBJECTIVE: Selective uptake of high density lipoprotein (HDL) cholesteryl ester (CE) is considered as the major source of cholesterol for production of steroids in the adrenal gland in rodents. As paraoxonase 1 (PON1) is an HDL-associated lipo-lactonase that has been shown to increase binding of HDL to macrophages, we used PON1 knock-out (PON1KO) mice to test the possible role of PON1 in corticosterone (CS) biosynthesis. METHODS AND RESULTS: PON1 deficiency was associated with reduced serum CS concentration. Adrenal glands obtained from PON1KO mice had significantly lower CE content compared to adrenals from C57Bl6 control mice. Binding of HDL obtained from PON1KO mice to human adrenocortical carcinoma cell line was found to be significantly lower than that of control HDL, and was associated with decreased CS biosynthesis. Addition of purified PON1 to HDL from PON1KO mice increased HDL binding and CS synthesis. Furthermore, the expression of the HDL receptor, SR-BI, protein and mRNA, was reduced in adrenals from PON1KO mice compared to control mice. When challenged with low salt diet, PON1KO mice demonstrated an increase in adrenal SR-BI gene expression and in serum corticosterone which reached levels similar to those obtained in control mice. CONCLUSION: PON1 regulates adrenal CS biosynthesis at two levels: (a) via an accessory role in HDL binding properties, and (b) a supportive role in SR-BI expression and CE supply to the cells.

Urokinase plasminogen activator upregulates paraoxonase 2 expression in macrophages via an NADPH oxidase-dependent mechanism.

OBJECTIVE: Macrophage foam cells are characterized by increased oxidative stress. Macrophage urokinase plasminogen activator (uPA) was shown to contribute to atherosclerosis progression. We hypothesized that uPA atherogenicity is related to its ability to increase macrophage oxidative stress. Increased macrophage oxidative stress in turn was shown to enhance PON2 expression. In the present study we investigated the effect of uPA on macrophage PON2 expression in relation to cellular oxidative stress. METHODS AND RESULTS: uPA increased PON2 expression in THP-1 macrophages in a dose-dependent manner. This effect required uPA/uPAR interaction and was abolished by cell treatment with antioxidants. uPA increased macrophage oxidative stress, measured by increased lipid peroxides, reactive oxygen species formation, superoxide anion release, and cell-mediated LDL oxidation. These effects were related to uPA-mediated activation of NADPH oxidase, and could not be reproduced in mouse peritoneal macrophages (MPM) harvested from p47(phox)-/- mice, suggesting a causal relationship between NADPH oxidase activation and the effects of uPA on macrophage oxidative stress and PON2 expression. Finally, MPM from PON2(-/-) mice were more susceptible to uPA-induced cellular oxidative stress than wild-type MPM, suggesting that PON2 protects against uPA-stimulated macrophage oxidative stress. CONCLUSIONS: Upregulation of macrophage PON2 may provide a compensatory protective mechanism against uPA-stimulation of macrophage oxidative stress during atherogenesis.

Paraoxonase 1 (PON1) attenuates diabetes development in mice through its antioxidative properties.

Paraoxonase 1 (PON1) is a lipo-lactonase which is associated with HDL and possesses antioxidative properties. Diabetes is characterized by increased oxidative stress and by decreased PON1 activity. We aimed to analyze whether oxidative status and PON1 levels in mouse sera and macrophages could affect streptozotocin (STZ)-induced diabetes development. We have used two models of mice under low oxidative stress: STZ-injected apolipoprotein E-deficient mice supplemented with the antioxidant vitamin E, and P47(phox) knockout mice. In both mice models the decreased serum basal oxidative stress, was associated with a decreased rate of diabetes development, compared with control STZ-injected apolipoprotein E-deficient mice or with C57BL mice respectively. These data suggest that oxidative stress accelerates diabetes development. Next, we analyzed the effect of PON1 on macrophage oxidative stress and on diabetes development in STZ-injected C57BL mice, PON1 knockout mice, and PON1 transgenic mice. PON1 overexpression was associated with decreased diabetes-induced macrophage oxidative stress, decreased diabetes development, and decreased mortality, in comparison to C57BL mice, and even more so when compared to PON1KO mice. We thus concluded that on increasing PON1 expression in mice, diabetes development is attenuated, a phenomenon which could be attributed to the antioxidative properties of PON1, as decrement of oxidative stress significantly attenuated STZ-induced diabetes development.

Macrophage paraoxonase 2 (PON2) expression is upregulated by unesterified cholesterol through activation of the phosphatidylinositol 3-kinase (PI3K) pathway.

Advanced atherosclerotic lesions are characterized by a progressive increase in the unesterified cholesterol (UC) content and a decrease in its cholesteryl ester (CE) content. In the present study, we examined mechanisms involved in the effect of UC and CE on the expression of paraoxonase 2 (PON2) in macrophages. J774A.1 macrophages were enriched with CE or UC by incubation for 14-48 h with 50 microg acetylated low-density lipoprotein in the absence or presence of the acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor 58035 (50 microg/ml), respectively. Macrophage PON2 mRNA expression, protein abundance and activity were increased only in the UC-enriched cells. In UC-enriched cells, inhibition of phosphatidylinositol 3-kinase (PI(3)K; using wortmannin or LY294002) attenuated the increase in PON2 mRNA expression by 50%, compared to untreated cells. In addition, we evidenced an increased phosphorylation of Akt in UC-enriched cells. Thus, we conclude from our data that macrophage PON2 expression is upregulated in UC-enriched macrophages through activation of the PI(3)K signal pathway.

Macrophage paraoxonase 2 (PON2) expression is up-regulated by pomegranate juice phenolic anti-oxidants via PPAR gamma and AP-1 pathway activation.

Paraoxonase 2 (PON2), a member of the paraoxonase gene family, was shown to protect macrophages against oxidative stress. Pomegranate juice (PJ), which contains potent polyphenol anti-oxidants, exhibits similar effects. We questioned possible association between PJ polyphenolics, macrophage oxidative stress, and cellular PON2 expression, in relation to the activation of specific PON2 transcription factors. Incubation of J774A.1 macrophages with PJ (0-50 microM of total polyphenols) dose-dependently increased expression (mRNA, protein) and activity and reduced macrophage oxidative status. These effects could be attributed to the PJ unique polyphenols, punicalagin and gallic acid. PJ polyphenol-induced up-regulation of PON2 was inhibited by 40% upon using the PPAR gamma inhibitor GW9662 (50 microM). Accordingly, the PPAR gamma ligand, rosiglitazone, dose-dependently stimulated macrophage PON2 expression, by up to 80%. Inhibition of AP-1 activation with SP600125, attenuated PJ-induced up-regulation of PON2 by 40%. Similarly, incubation of macrophages with PJ polyphenols in the presence of GW9662 or SP600125, significantly reduced their capacity to protect the cells against oxidative stress. We conclude that the anti-oxidative characteristics of PJ unique phenolics punicalagin and gallic acid could be related, at least in part, to their stimulatory effect on macrophage PON2 expression, a phenomenon which was shown to be associated with activation of the transcription factors PAPR gamma and AP-1.

A biphasic U-shape effect of cellular oxidative stress on the macrophage anti-oxidant paraoxonase 2 (PON2) enzymatic activity.

Expression of macrophage paraoxonase 2 (PON2), a cellular lactonase with anti-oxidant and anti-atherogenic properties, was shown to be upregulated under high oxidative stress. The aim of the present study was to analyze the relationship between the extent of cellular oxidative stress in J774A.1 macrophage and PON2 lactonase activity under various levels of oxidation, obtained by cell incubation with either anti-oxidants or oxidants. PON2 activity exhibited a U-shape response curve. In the oxidative stress range below that of control untreated cells, PON2 activity decreased upon increasing macrophage oxidative state, whereas in the range over that of control untreated cells, PON2 activity increased. The biphasic effect of oxidative stress on macrophage PON2 activity could be related to PON2 inactivation (decreased enzymatic activity) under oxidative stress induction at its low range, whereas at high range of oxidative stress, macrophage anti-oxidant compensatory mechanism up-regulates PON2 (increased protein expression), in order to cope with oxidative burden.

Paraoxonase 2 (PON2) expression is upregulated via a reduced-nicotinamide-adenine-dinucleotide-phosphate (NADPH)-oxidase-dependent mechanism during monocytes differentiation into macrophages.

Paraoxonase 2 (PON2) is a member of the paraoxonases gene family. PON2 is ubiquitously present in cells, including macrophages, and it was shown to protect against cellular oxidative stress. The aim of the present study was to analyze mechanisms involved in PON2 expression during monocyte/macrophage differentiation. PON2 expression was analyzed in vitro in THP-1 cells differentiated with 1alpha,25-dihydroxyvitamin D3 and in vivo in mouse peritoneal macrophages (MPM) isolated at increasing time intervals after intraperitoneal thioglycollate injection. PON2 expression (mRNA and protein) and activity gradually increased during monocyte/macrophage differentiation, up to five fold and eight fold in vitro and in vivo, respectively. This effect was associated with a gradual increase in cellular superoxide anion production. Supplementation of vitamin E to Balb/C mice inhibited the reduced nicotinamide adenine dinuleotide phosphate (NADPH)-oxidase-dependent increase in cellular superoxide anion production by 50% and down-regulated PON2 mRNA expression and activity by 30 and 60%, respectively. Furthermore, PON2 expression was lower by nine fold in MPM isolated from P47(phox-/-) (inactive NADPH oxidase) mice, in comparison to MPM from control mice. PON2 expression was found to be regulated, at least in part, by the transcription factor AP-1, as suggested by decreased JDP2 (AP-1 repressor) protein expression in the nucleus and by decreased PON2 expression in the presence of a Jun N-terminal kinase inhibitor (SP600125). The present study demonstrates, for the first time, that PON2 expression increases in monocytes during their maturation into macrophage as a result of NADPH-oxidase activation, and this process is partly regulated by the transcription factor AP-1. PON2 stimulation may represent a compensatory mechanism against the increase in cellular superoxide anion production and atherogenesis.

Cell-induced copper ion-mediated low density lipoprotein oxidation increases during in vivo monocyte-to-macrophage differentiation.

Macrophage activation is associated with the production and release of reactive oxygen species (ROS), which are capable of mediating oxidative modification of low-density lipoprotein (LDL). In the present study we questioned whether cellular capacity to oxidize LDL increases during in vivo monocyte/macrophage maturation. We developed a novel model for macrophage maturation in vivo using mouse peritoneal macrophages (MPMs) harvested at increasing intervals after intraperitoneal thioglycollate injection. Macrophage maturation was evidenced by a progressive increase in cellular size, density, granulation, and expression of cell surface markers CD11b and CD36, and by a gradual decrement in myeloperoxidase activity. Cellular capacity to stimulate copper ion-mediated oxidation of LDL increased gradually by up to 2-fold during in vivo macrophage maturation in Balb/C mice, similar to the pattern observed during 1,25-dihydroxyvitamin D3-induced in vitro differentiation of the PLB-985 cell line. These effects were attributed to a gradual increase in production of ROS by up to 9-fold. The mechanism for the increase in cellular oxidative stress during macrophage maturation could be related, at least in part, to NADPH oxidase activation, as demonstrated by a gradual increase over time in p47phox expression (mRNA and protein) and in its translocation to the plasma membrane. In conclusion, in vivo monocyte-to-macrophage differentiation is associated with increased cell capacity to oxidize LDL, which may represent a protective mechanism for rapid removal of atherogenic LDL from extracellular spaces in the arterial wall.